The mammalian reproductive system is physiologically and anatomically differentiated between the sexes. These sex differences range from the prevalence of certain health disorders of reproductive function to differences in normal reproductive physiology and reproductive neural circuits. In rodents, the anteroventral periventricular nucleus (AVPV) of the hypothalamus, a region important for gonadotropin secretion, is a well- known sexually dimorphic region, being larger and having more cells in females than males. Certain neuronal populations in the AVPV are also sexually dimorphic, such as Kiss1 cells, with females having many more AVPV Kiss1 neurons than males. The Kiss1 gene encodes kisspeptin, a neuropeptide which has recently been implicated in fertility and puberty in many species, including humans. The sexual differentiation of Kiss1 neurons in the AVPV may underlie the ability of adult females, but not males, to produce the preovulatory gonadotropin surge, and may also relate to sex differences in puberty onset, which typically occurs earlier in females than males. Like many other sexually dimorphic brain populations, the Kiss1 system is sexually differentiated under the influence of the sex steroid hormone milieu during the postnatal "critical period" of development. However, much is still unknown about the specific hormonal and molecular mechanisms underlying the sex steroid-induced sexual differentiation of the brain and Kiss1 system. Sex steroids may regulate sexual differentiation of neuronal populations, including Kiss1 cells, via several mechanisms, including differential cell death (apoptosis), neurogenesis, and/or epigenetic alterations. Of these mechanisms, apoptosis is the most common process implicated in sexual differentiation of the brain. Importantly, the pro- apoptotic protein BAX is required for the sexual differentiation of the AVPV region as a whole, suggesting it may also be involved in controlling the development of the sexually dimorphic Kiss1 system. This proposal addresses several important questions regarding the mechanisms of sexual differentiation of neuroendocrine reproductive circuits. Aim 1 analyzes the development and phenotypic identity of the sexually differentiated Kiss1 system. Aim 2 studies the specific contributions of apoptosis, neurogenesis, and epigenetics to the sexual differentiation of Kiss1 neurons. Lastly, due to the importance of the pro-apoptotic gene Bax in the sexual differentiation of the AVPV (and other brain regions), Aim 3 investigates the specific molecular and cellular mechanisms by which sex steroids induce Bax-mediated apoptosis in cultured hypothalamic neurons. Overall, this proposal will provide a better understanding of the development of Kiss1 neurons, as well as provide insight into the molecular and cellular mechanisms by which sex steroids control sexual differentiation of the brain. Because kisspeptin is essential for governing puberty and reproduction, these studies will also provide novel insight into potential mechanisms underlying sexually differentiated reproductive disorders, such as hypogonadotropic hypogonadism (more common in men) and precocious puberty (more common in girls). PUBLIC HEALTH RELEVANCE: Several facets of reproduction are sexually differentiated, including the onset of puberty and the ability to generate a preovulatory luteinizing hormone surge, as well as a number of reproductive diseases and disorders, such as idiopathic hypogonadotropic hypogonadism (more common in men), precocious puberty (more prevalent in girls), and constitutional delayed puberty (more common in boys). This research proposal investigates the underlying hormonal, cellular, and molecular mechanisms contributing to the sexual differentiation of kisspeptin neurons, which have been implicated in regulating puberty and fertility and may therefore be involved in the etiology of sexually dimorphic reproductive disorders. This work will contribute to our understanding of how developmental sex steroids influence the sexual differentiation of reproductive neural circuits and will ultimately provide insight into developmental regulation of several reproductive disorders.